The CD28-mediated costimulatory signal plays a pivotal role in the outcome of many immune responses including cytolytic responses in tumor and autoimmune diseases. Depending on the primary stimulation, CD28 can initiate multiple intracellular signaling pathways including a pathway that is insensitive to immunosuppressive drug, Cyclosporin A (CsA). This CsA- insensitive pathway is believed to be involved in graft-vs-host disease (GVHD) during allogeneic bone marrow transplantation. Our current objectives focus on three areas: (1) characterization of the CsA-resistant rapamycin-sensitive pathway of T cell activation; (2) examination of the physiological significance of this pathway; and (3) the effect of aging on the rapamycin-sensitive pathway. Our recent work has demonstrated that the immunosuppressive drug rapamycin selectively affects the CsA-resistant pathway. Our initial studies have focused on the mechanism of activation of the IL-2 gene in a CsA-resistant manner. We found that the effects of rapamycin on the IL-2 expression was due to alteration in IL-2 mRNA stability. More recently, we have also shown that activation of T cells by IL-12 is resistant to CsA, but sensitive to rapamycin. As the intracellular target of rapamycin is mTOR, we are investigating the mechanism of activation of mTOR by CD28 costimulation, and the role of mTOR in IL-2 mRNA stability. To examine the physiological role of this resistant pathway, we are currently examining the role of this pathway in the differentiation of T cells as well as allogeneic mixed lymphocyte reactions (MLR), an in vitro model of GVHD. We have initiated studies to dissect the CsA- sensitive component of the allogeneic mixed lymphocyte reaction from the CsA-resistant one. We believe that further elucidation of this signaling pathway may assist in the identification of novel therapeutics to prevent GVHD.